Massively parallel in vivo CRISPR screening identifies RNF20/40 as epigenetic regulators of cardiomyocyte maturation

VanDusen, Nathan J.; Lee, Julianna Y.; Gu, Weiliang; Butler, Catalina E.; Sethi, Isha; Zheng, Yanjiang; King, Justin S.; Zhou, Pingzhu; Suo, Shengbao; Guo, Yuxuan; Ma, Qing; Yuan, Guo‐Cheng; Pu, William William

doi:10.1038/s41467-021-24743-z

Cited by 40 publications

(31 citation statements)

References 72 publications

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“…The functions and gene targets of GATA6 are partially redundant with GATA4 35 – 37 . To determine whether ERRγ and GATA6 also shared targets, we conducted an intersection analysis with GATA6 ChIP-seq in hiPSC-CMs 18 and our ERRγ ChIP-seq datasets.…”

Section: Resultsmentioning

confidence: 99%

The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

et al. 2022

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Estrogen-related receptors (ERR) α and γ were shown recently to serve as regulators of cardiac maturation, yet the underlying mechanisms have not been delineated. Herein, we find that ERR signaling is necessary for induction of genes involved in mitochondrial and cardiac-specific contractile processes during human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) differentiation. Genomic interrogation studies demonstrate that ERRγ occupies many cardiomyocyte enhancers/super-enhancers, often co-localizing with the cardiogenic factor GATA4. ERRγ interacts with GATA4 to cooperatively activate transcription of targets involved in cardiomyocyte-specific processes such as contractile function, whereas ERRγ-mediated control of metabolic genes occurs independent of GATA4. Both mechanisms require the transcriptional coregulator PGC-1α. A disease-causing GATA4 mutation is shown to diminish PGC-1α/ERR/GATA4 cooperativity and expression of ERR target genes are downregulated in human heart failure samples suggesting that dysregulation of this circuitry may contribute to congenital and acquired forms of heart failure.

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Section: Resultsmentioning

confidence: 99%

The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

et al. 2022

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“…Furthermore, extensive studies have been conducted on the mechanisms by which H3K4 and H3K27 methylations impact heart development 11,14,15 . More recently, a critical role for histone H2B ubiquitylation (H2Bub) was demonstrated in cardiac development 16,17 . Knockdown of the Xenopus H2Bub E3 ligases, rnf20 (Ring finger protein 20) and rnf40 (Ring finger protein 40) 18 , beginning at the two-cell stage, influences cardiac looping and left-right (LR) asymmetry during early embryonic development 16 .…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, the results of an in vivo CRISPR/Cas9 genetic screen suggested that RNF20/40 is involved in cardiomyocyte (CM) maturation via H2Bub deposition 17 . Thus, RNF20 appears to be relevant to both CHD and CM maturation 16,19 . Nevertheless, the molecular function of RNF20 during temporal development of mammalian heart remains uncharacterized.…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, the results of an in vivo CRISPR/Cas9 genetic screen suggested that RNF20/40 is involved in cardiomyocyte (CM) maturation via H2Bub deposition 18 . Thus, RNF20 appears to be relevant to both CHD and CM maturation 17,20 . The addition of a single ubiquitin on H2B disrupts local and higher-order structures of compacted chromatin 21,22 .…”

Section: Introductionmentioning

confidence: 99%

“…Knockdown of the Xenopus H2Bub E3 ligases, rnf20 (Ring finger protein 20) and rnf40 (Ring finger protein 40) 19 , beginning at the two-cell stage, influences cardiac looping and left-right (LR) asymmetry during early embryonic development 17 . Furthermore, the results of an in vivo CRISPR/Cas9 genetic screen suggested that RNF20/40 is involved in cardiomyocyte (CM) maturation via H2Bub deposition 18 . Thus, RNF20 appears to be relevant to both CHD and CM maturation 17,20 .…”

Section: Introductionmentioning

confidence: 99%

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Chromatin state transition underlies the temporal changes in gene expression during cardiomyocyte maturation

Lin

Chang

Tseng

et al. 2021

Preprint

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Congenital heart disease (CHD) is often rooted in aberrant gene expression during heart development. As cells commit to a specific lineage during development, chromatin dynamics and developmental plasticity generally become more limited. However, it remains unclear how differentiated cardiomyocytes (CMs) undergo morphological and functional adaptations to the postnatal environment during the process of CM maturation. We sought to investigate the regulatory mechanisms that control postnatal cardiac gene networks. A time-series transcriptomic analysis of postnatal hearts revealed an integrated, time-ordered transcriptional network that regulates CM maturation. Remarkably, depletion of histone H2B ubiquitin ligase RNF20 after formation of the four-chamber heart disrupted these highly coordinated gene networks. As such, its ablation caused early-onset cardiomyopathy, a phenotype reminiscent of CHD. Furthermore, the dynamic modulation of chromatin accessibility by RNF20 during CM maturation was necessary for the operative binding of cardiac transcription factors that drive transcriptional gene networks. Together, our results reveal how epigenetic-mediated chromatin state transitions modulate time-ordered gene expression for CM maturation.

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Engineered Cardiac Tissues as a Platform for CRISPR‐Based Mitogen Discovery

DeLuca,

Strash,

Chen

et al. 2024

Adv Healthcare Materials

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Improved understanding of cardiomyocyte (CM) cell cycle regulation may allow researchers to stimulate pro‐regenerative effects in injured hearts or promote maturation of human stem cell‐derived CMs. Gene therapies, in particular, hold promise to induce controlled proliferation of endogenous or transplanted CMs via transient activation of mitogenic processes. Methods to identify and characterize candidate cardiac mitogens in vitro can accelerate translational efforts and contribute to the understanding of the complex regulatory landscape of CM proliferation and postnatal maturation. In this study, A CRISPR knockout‐based screening strategy using in vitro neonatal rat ventricular myocyte (NRVM) monolayers is established, followed by candidate mitogen validation in mature 3‐D engineered cardiac tissues (ECTs). This screen identified knockout of the purine metabolism enzyme adenosine deaminase (ADA‐KO) as an effective pro‐mitogenic stimulus. RNA‐sequencing of ECTs further reveals increased pentose phosphate pathway (PPP) activity as the primary driver of ADA‐KO‐induced CM cycling. Inhibition of the pathway's rate limiting enzyme, glucose‐6‐phosphate dehydrogenase (G6PD), prevented ADA‐KO induced CM cycling, while increasing PPP activity via G6PD overexpression increased CM cycling. Together, this study demonstrates the development and application of a genetic/tissue engineering platform for in vitro discovery and validation of new candidate mitogens affecting regenerative or maturation states of cardiomyocytes.

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Massively parallel in vivo CRISPR screening identifies RNF20/40 as epigenetic regulators of cardiomyocyte maturation

Cited by 40 publications

References 72 publications

The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

Chromatin state transition underlies the temporal changes in gene expression during cardiomyocyte maturation

Engineered Cardiac Tissues as a Platform for CRISPR‐Based Mitogen Discovery

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